Reducing morbidity in surgical resections: Third-harmonic generation microscopy a

描述（由申请方提供）：神经意外损伤是肿瘤手术切除期间以及其他手术中发病的主要原因之一。在手术过程中，神经纤维很难与周围组织区分开来，这使得外科医生很难仔细工作并避免其受伤。从广义上讲，所提出的工作的目标是建立三次谐波产生（THG）显微镜作为一种可视化的方法，具有高灵敏度，神经纤维术中。我们以前已经表明，THG提供了一种高灵敏度的方法来成像髓鞘在啮齿动物的中枢神经系统，并已使用此在脊髓中的髓鞘在体内成像。我们最近收集的初步数据表明，这种方法也可以突出周围神经。最终，这种成像方式可以让外科医生可视化神经纤维，然后在它们附近仔细工作，从而防止与神经损伤相关的手术并发症。我们确定了完成这项工作所需的三个具体目标。首先，我们建立在我们的初步数据，并确定最佳的一组激光和成像参数，以可视化有髓鞘的周围神经束与THG成像。第二，我们通过比较啮齿动物中使用和不使用这种新的成像方式进行前列腺切除手术的结果，评估THG成像对保留神经的有效性。在这个目标中，我们进一步建立在以前的结果，并检查不仅如何以及神经被保存时，可视化与THG，而且如何完全提取肿瘤时，他们与双光子激发荧光（2 PEF）显微镜成像。至关重要的是，THG成像所需的激光和成像系统与2 PEF成像完全兼容，允许这两种模式协同工作，以帮助外科医生最大限度地切除肿瘤并最大限度地减少神经损伤。两者结合起来，这两个目标将建立我们的神经成像方法的实用性，但在手术中使用， 室内环境显然需要开发一种比我们目前的显微镜更轻便的仪器。为此，在第三个目标中，我们设计了一种内窥镜，该内窥镜针对THG（以及2 PEF）成像进行了优化，并可用于人类腹腔镜手术（如许多前列腺癌切除术）。总之，这项工作将建立THG作为一种可行的成像方式用于手术的效用，并将开发可用于手术室的第一代仪器。如果成功，这项工作可以大大降低切除手术的术后发病率-特别是在关键神经靠近切除部位的情况下，例如前列腺-从而改善患者的预后。

DESCRIPTION (provided by applicant): Unintentional injury to nerves is one of the primary causes of morbidity during surgical resection of tumors as well as in other surgeries. Nerve fibers are difficult to distinguish from the surrounding tissues during surgery, making it difficul for surgeons to carefully work around and avoid their injury. Broadly speaking, the goal of the proposed work is to establish third-harmonic generation (THG) microscopy as an approach to visualize, with high sensitivity, nerve fibers intraoperatively. We have previously shown that THG provides a high-sensitivity approach to imaging myelin in the central nervous system of rodents and have used this for in vivo imaging of myelin in the spinal cord. We recently collected preliminary data that shows this approach can highlight peripheral nerves as well. Ultimately, this imaging modality could allow the surgeon to visualize nerve fibers and then work carefully near them, thereby preventing surgical complications associated with nerve injury. We identify three Specific Aims necessary to complete this work. First, we build on our preliminary data and identify the best set of laser and imaging parameters to visualize myelinated peripheral nerve bundles with THG imaging. Second, we assess the effectiveness of THG imaging to spare nerves by comparing the outcome of prostate resection surgeries in rodents done with and without this novel imaging modality. In this Aim, we further build on previous results and examine not just how well nerves are spared when visualized with THG, but also how completely tumors are extracted when they are imaged with two-photon excited fluorescence (2PEF) microscopy. Critically, the laser and imaging systems necessary for THG imaging are completely compatible with 2PEF imaging, allowing these two modalities to work in concert to help a surgeon both maximize tumor extraction and minimize nerve damage. Taken together, these two aims will establish the utility of our approach to nerve imaging, but use in an operating room environment will clearly require development of a less-unwieldy instrument than our current microscopes. Toward this end, in the third Aim, we design an endoscope that is optimized for THG (as well as 2PEF) imaging and could be used in laparoscopic surgeries (such as many prostate cancer resections) in humans. Taken together, this work will establish the utility of THG as a viable imaging modality for use in surgery and will develop the first-generation instrument that could be used in an operating room. If successful, this work could dramatically reduce the post-surgical morbidity of resection surgeries - especially in cases where critical nerves are close to the resection site, such as in the prostate - and therefore improve patient outcomes.